Very large scale immobilized polymer synthesis

Abstract

A synthetic strategy for the creation of large scale chemical diversity. Solid-phase chemistry, photolabile protecting groups, and photolithography are used to achieve light-directed spatially-addressable parallel chemical synthesis. Binary masking techniques are utilized in one embodiment. A reactor system, photoremovable protecting groups, and improved data collection and handling techniques are also disclosed. A technique for screening linker molecules is also provided.

1 . A method of detecting hybridization between biological polymers, comprising the acts of: 
 providing a substrate having a surface including at least one biological polymer and at least one fluorescent label associated with the biological polymer;    generating an excitation laser beam;    scanning said laser beam relative to said surface; collecting fluorescent radiation responsive to said laser beam using optics;    detecting said collected fluorescent radiation; and    autofocusing to bring into focus with respect to said optics at least a portion of said surface including the biological polymer.    
     
     
         2 - 62 . (canceled)  
     
     
         63 . A method for synthesizing a plurality of biopolymers on the surface of a support, said method comprising: (a) placing said support into a reaction chamber and applying to said surface said biopolymers or precursors of said biopolymers, (b) removing said support from said reaction chamber and placing said support into a flow chamber, (c) introducing a liquid reagent for conducting said synthesis into said flow chamber, (d) removing said liquid reagent from said flow chamber wherein the pressure in said chamber is maintained substantially atmospheric during said removing. (e) removing said support from said flow chamber and (f) repeating steps (a)-(e) to form said plurality of biopolymers on the surface of said support.  
     
     
         64 . A method according to  claim 63  wherein liquid reagent is removed from said flow chamber under vacuum.  
     
     
         65 . A method according to  claim 63  wherein liquid reagent is removed from said flow chamber by simultaneously venting and applying a vacuum to said flow chamber.  
     
     
         66 . A method according to  claim 89  wherein said venting and said applying a vacuum are carried out at opposite ends of said flow chamber.  
     
     
         67 . A method according to  claim 63  wherein said method further comprises holding said liquid reagent in said flow chamber for a predetermined period of time.  
     
     
         68 . A method according to  claim 63  wherein said support is glass.  
     
     
         69 . A method according to  claim 63  further comprising introducing a pressurized inert gas into said flow chamber after step (c) and simultaneously evacuating said flow chamber.  
     
     
         70 . A method according to  claim 63  wherein said biopolymers are polynucleotides.  
     
     
         71 . A method according to  claim 63  wherein said liquid reagent for conducting said synthesis comprises an oxidizing agent or an agent for removing a protecting group.  
     
     
         72 . A method according to  claim 63  wherein said biopolymers are synthesized on said surface in multiple arrays and said support is subsequently diced into individual arrays of biopolymers on a support.  
     
     
         73 . A method according to  claim 72  further comprising exposing the array to a sample and reading the array.  
     
     
         74 . A method according to  claim 73  comprising forwarding data representing a result obtained from a reading of the array.  
     
     
         75 . A method according to  claim 74  wherein the data is transmitted to a remote location.  
     
     
         76 . A method according to  claim 75  comprising receiving data representing a result of an interrogation obtained by the reading of the array.  
     
     
         77 . A method for synthesizing an array of biopolymers on the surface of a support wherein said synthesis comprises a plurality of monomer additions, said method comprising after each of said monomer additions: (a) placing said support into a flow chamber, (c) introducing a liquid reagent for conducting said synthesis into said flow chamber, (d) removing said reagent from said flow chamber by simultaneously venting said chamber and applying a vacuum to the interior of said chamber, (e) removing said support from said flow chamber and (f) repeating steps (a)-(e) to form said plurality of biopolymers on the surface of said support.  
     
     
         78 . A method according to  claim 77  wherein said venting and said applying a vacuum are carried out at opposite ends of said flow chamber.  
     
     
         79 . A method according to  claim 77  wherein said method further comprises holding said liquid reagent in said flow chamber for a predetermined period of time.  
     
     
         80 . A method according to  claim 77  wherein said support is glass.  
     
     
         81 . A method according to  claim 77  further comprising introducing a pressurized inert gas into said flow chamber after step (d) and simultaneously evacuating said flow chamber.  
     
     
         82 . A method according to  claim 77  wherein said biopolymers are polynucleotides.  
     
     
         83 . A method according to  claim 77  wherein said liquid reagent for conducting said synthesis is an oxidizing agent or an agent foi removing a protecting group.  
     
     
         84 . A method according to  claim 77  wherein said biopolymers are synthesized on said surface in multiple arrays and said support is subsequently diced into individual arrays of biopolymers on a support.  
     
     
         85 . A flow cell assembly for conducting at least one reaction in the synthesis of an array of biopolymers on the surface of a support, said flow cell comprising: (a) a flow cell chamber, (b) a manifold in fluid communication with said chamber, said manifold comprising at least a wash reagent inlet, an inlet for a reagent for conducting a step of said synthesis, and a vent, and (c) a vacuum source in fluid communication with said flow cell chamber.  
     
     
         86 . A flow cell assembly according to  claim 85  further comprising a fluid level sensor and a controller for controlling said inlets, said vent and said vacuum source.  
     
     
         87 . A flow cell assembly according to  claim 85  further comprising a gas inlet.  
     
     
         88 . An apparatus for synthesizing an array of biopolymers on the surface of a support, said apparatus comprising: (a) one or more flow cell assemblies of  claim 85 , (b) one or more fluid dispensing stations in fluid communication with one or more of said plurality of flow cell assemblies, (c) a station for monomer addition to said surface of said support, and (d) a mechanism for moving a support to and from said station for monomer addition and a flow cell and from one flow cell to another flow cell.  
     
     
         89 . An apparatus according to  claim 88  further comprising a controller for controlling the movement of said mechanism.  
     
     
         90 . An apparatus according to  claim 88  wherein said mechanism is a robotic arm.  
     
     
         91 . A method comprising using an array, prepared by an apparatus according to  claim 88 , by exposing the array to a sample and reading the array.  
     
     
         92 . A method according to  claim 91  comprising forwarding data representing a result obtained from a reading of the array.  
     
     
         93 . A method according to  claim 92  wherein the data is transmitted to a remote location.  
     
     
         94 . A method according to  claim 93  comprising receiving data representing a result of an interrogation obtained by the reading of the array.